Human embryonic stem cells (hESCs) can be proliferated indefinitely while retaining a normal karyotype, and can be used in vitro to study the earliest stages of differentiation. While in vitro studies (drug testing, target identification) may ultimately be medically more valuable than in vivo applications, it is the latter that has captured our imagination. The decision to fund research on hESCs opens up unprecedented opportunities. Public funding takes the research out of the private sector, where it has been motivated solely by economic necessities, into the public sector, where it can be performed without the driving force of the marketplace. The course proposed in this application, "CHOC Human Embryonic Stem Cell Culture Training Course", is a continuation of our current successful course which provides hands-on training for investigators to learn how to culture, manipulate, and differentiate hESCs in vitro. This course brings together some of the leading experts on hESC technology and through comparative approaches, effectively trains students in the successful culture, maintenance and manipulation of hESCs using lines WA01 and WA09 from the NIH hESC Registry. A major long term goal for this course has been to share and improve standard protocols. Specifically, this course aims are: 1) Demonstration of proper growth conditions for cells; 2) Proper freeze-thaw cycling and preparation of cell passages; 3) Use of co-culture techniques; 4) Use of cell separation procedures; 5) Review of basic good laboratory practices for use of human biological materials; 6) Development of protocols that support the characterization of embryonic stem cells; 7) Application of standard research protocols for directed differentiation of embryonic stem cells; 8) Employment of standard detection methods for infectious organisms or other contaminants; and 9) Cross-training of techniques used in other applications of stem cell biology. These aims are accomplished by offering intensive 10-day "immersion" courses to 12 participants chosen for their outstanding research potential and commitment to institute hESC research in their own laboratories. In addition, the future courses will each conclude with a day-long symposium, "Cellular Therapies in Pediatric Diseases", designed to highlight the clinical goals and obstacles for hESC research. This symposium, which focuses primarily on pediatric diseases, will be open to the general public as well as biomedical researchers and clinicians and, for the latter, will provide Continuing Medical Education credits. The courses are held annually in the Spring at the National Human Neural Stem Cell Resource of the Neuroscience Laboratories at the Children's Hospital of Orange County Research Institute. [unreadable] [unreadable] [unreadable]